Alv eolar type I (AT1) cells are believed to be derived from type II (AT2) cells in vivo during normal turnover of the alveolar epithelium and following injury through a process termed 'transdifferentiation'. However, the molecular mechanisms that mediate transitions between alveolar epithelial cell (AEC) phenotypes, and that regulate cellular differentiation within adult alveolar epithelium, remain almost entirely unknown. We have developed an in vitro culture model of AEC transdifferentiation and reverse transdifferentiation that provides an excellent tool with which to investigate transcriptional pathways that regulate AEC differentiation. FoxP2 is the only member of the Fox family of transcription factors (TF) that is exclusively expressed in distal lung epithelium. FoxP2 regulates transcriptional activity of several lung-enriched genes and appears necessary for normal AEC differentiation in the early postnatal period. The goal of this proposal is to investigate the role of FoxP2 in mediating transitions between AT2 and AT1 cell phenotypes through its effects on expression of AT2 and AT1 cell differentiation-related genes. We hypothesize that: 1) changes in the levels/activity of FoxP2 modulate cell type-specific gene expression during transitions between AT2 and AT1 cell phenotypes in adult alveolar epithelium, and 2) dynamic interplay between FoxP2 and other TF that are differentially expressed in AT2 and AT1 cells determines the state of AEC differentiation. We will use our well-characterized model of AEC differentiation in vitro, in conjunction with freshly isolated AT2 and AT1 cells and our expertise in characterizing AEC differentiation, to test these hypotheses by addressing the following Specific Aims: 1) investigate regulation of expression of FoxP2 and novel FoxP2-interacting TF in AEC, 2) determine individual and combinatorial roles of FoxP2 and other TF in regulating expression of AT2 and AT1 cell-restricted genes, and 3) investigate functional consequences of under- and over-expression of FoxP2 on AEC phenotype. We will evaluate expression of FoxP2 in isolated cells and in AEC in vitro and in situ, identify putative FoxP2-interacting factors in AT1 and AT2 cells, characterize interactions of FoxP2 with, and promoter occupancy of, AT2 (SP-C) and AT1 (AQP5) cell-specific genes and identified interacting TF, and assess effects of modulating FoxP2 expression on AEC phenotype. Knowledge of transcriptional programs that mediate transitions between AT2 and AT1 cell phenotypes, and that lead to distinct patterns of gene expression in differentiated AT2 and AT1 cells, are essential for understanding maintenance of alveolar epithelium and restoration of its cellular architecture following injury.